Molecular oxygen is an essential nutrient for nonfacultative aerobic organisms, including humans. Oxygen, although essential for aerobic metabolism, can be converted to poisonous metabolites, such as superoxide anion and hydrogen peroxide, collectively known as reactive oxygen species. Excessive concentrations of various forms of oxygen and of free radicals can have serious adverse effects on living systems, including the peroxidation of membrane lipids, the hydroxylation of nucleic acid bases, and the oxidation of sulfhydryl groups and other sensitive moieties in proteins. If uncontrolled, mutations and cell death result.
Biological antioxidants include well-defined naturally occurring metalloenzymes, such as superoxide dismutase (SOD), catalase (CAT), and selenium glutathione peroxidase, as well as the enzyme, phospholipid hydroperoxide glutathione peroxidase. A large number of diseases or degenerative processes are related to disorders with metalloenzymes involved in the detoxification of reactive oxygen species derived from dioxygen reduction. The role of these metalloenzymes has been demonstrated with animals under-expressing SOD or CAT enzymes. In addition, the induction of nitric oxide-dependent apoptosis in motor neurons by zinc-deficient superoxide dismutase has recently been shown (Estévez et al. (2000), Science, 286:2498-2500).
Reactive oxygen species are key executioners in Fas-induced hepatocyte apoptosis. (Malassagne et al., Gastroenterology 121:1451-1459 (2001)). Human Fas ligand is a polypeptide which has been reported by Nagata et al. to be a biological molecule which induces apoptosis of Fas-expressing cells (Takahashi, T. et al., International Immunology, vol, 6, 1567-1574, 1994). Human Fas ligand is a Type II membrane protein of TNF family with a molecular weight of about 40 kD. The extracellular domain of the human Fas ligand is highly homologous with the extracellular domain of rat Fas ligand (Suda, T. et al., Cell, vol. 75, 1169-1178, 1993) and mouse Fas ligand (Takahashi, T. et al., Cell, vol. 76, 969-976, 1994). The human Fas ligand recognizes not only the human Fas but also the mouse Fas to induce the apoptosis, and vice versa, the rat Fas ligand and the mouse Fas ligand also recognize the human Fas to induce the apoptosis.
Considerable research has been done on the mechanism of signal transduction in the cell upon the Fas-mediated apoptosis, and identification and cloning of the factor which interacts with the intracellular domain of the Fas, in particular, the region called “death domain” to transmit or block the signal have been reported. Possibility of the involvement of ICE (interleukin-1-converting enzyme)-related thiol proteases in the Fas-mediated apoptosis has also been indicated. Fas-Fas ligand interaction plays a major role in hepatoctye injury during viral hepatitis through the activation of caspases or through mitochondrial disruption.
Obstacles exist for the use of recombinant metalloenzymes in therapy including: solution instability, limited cellular accessibility, orally bioavailability, immunogenicity, short half-lives, cost of production and proteolytic digestion. These synthetic catalytic scavengers must be stable in physiological conditions and, in particular, the metal should be strictly inserted within the ligand to avoid any demetallation and trapping of the metal on by serum proteins. These synthetic catalytic scavengers must also be soluble in water at pH 7.0. It is desirable that the compositions are orally bioavailable. Avoiding synthetic molecules that lead to DNA cleavage is an additional concern.
Consequently, there is a need for new oral bioavailable synthetic transition metal complexes with the ability to scavenge reactive oxygen species derived from the non-controlled reduction of dioxygen. The need exists for providing low molecular weight, orally bioavailable water soluble metallophorphyrin derivatives able to scavenge reactive oxygen species.